title: Structural basis for eliciting a cytotoxic effect in HER2-overexpressing cancer cells via binding to the extracellular domain of HER2.
abstract: Human epidermal growth factor receptor-2 (HER2) is a receptor tyrosine kinase directly linked to the growth of malignancies from various origins and a validated target for monoclonal antibodies and kinase inhibitors. Utilizing a new approach with designed ankyrin repeat proteins (DARPins) as alternative binders, we show that binding of two DARPins connected by a short linker, one targeting extracellular subdomain I and the other subdomain IV, causes much stronger cytotoxic effects on the HER2-addicted breast cancer cell line BT474, surpassing the therapeutic antibody trastuzumab. We determined crystal structures of these DARPins in complex with the respective subdomains. Detailed models of the full-length receptor, constrained by its rigid domain structures and its membrane anchoring, explain how the bispecific DARPins connect two membrane-bound HER2 molecules, distorting them such that they cannot form signaling-competent dimers with any EGFR family member, preventing any kinase dimerization, and thus leading to a complete loss of signaling.
authors.affiliation: Department of Biochemistry, University of Zürich, 8057 Zürich, Switzerland.
authors.forename: Christian
authors.initials: C
authors.lastname: Jost
authors.forename: Johannes
authors.initials: J
authors.lastname: Schilling
authors.forename: Rastislav
authors.initials: R
authors.lastname: Tamaskovic
authors.forename: Martin
authors.initials: M
authors.lastname: Schwill
authors.forename: Annemarie
authors.initials: A
authors.lastname: Honegger
authors.forename: Andreas
authors.initials: A
authors.lastname: Plückthun
mesh_terms: D000818:Animals; D017089:Ankyrin Repeat; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology; D018931:Antineoplastic Agents, Hormonal / Q000737:chemistry / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D018360:Crystallography, X-Ray; D018604:Epitope Mapping; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000494:pharmacology*; D061987:Sf9 Cells; D015398:Signal Transduction; D018411:Spodoptera; D000068878:Trastuzumab
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D018931:Antineoplastic Agents, Hormonal; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D000068878:Trastuzumab; C508053:ERBB2 protein, human